PubMed Health⌕ Search

Biomedical subjects

W den Otter

Publications and source records attributed to W den Otter.

At least 19 recordsLinked to original sources
PubMed2002

Release of recombinant human interleukin-2 from dextran-based hydrogels.

In this study, the release of recombinant human interleukin-2 (rhIL-2) from methacrylated dextran (dex-MA) and (lactate-)hydroxyethyl methacrylated dextran (dex-(lactate-)HEMA) hydrogels with varying crosslink density was investigated. Hydrogels derived from dex-MA are stable under physiological conditions (pH 7 and 37 degrees C), whereas dex-HEMA and dex-lactate-HEMA hydrogels degrade due to the presence of hydrolytically sensitive esters in the crosslinks of the gels. The protein release profiles both the non-degradable and degradable dextran-based hydrogels showed that with increasing crosslink density of the gel, the release of rhIL-2 decreases. From dex-MA hydrogels with an initial water content above 70%, the rhIL-2 release followed Fickian diffusion, whereas from gels with an initial water content of 70% or lower the protein was fully entrapped in the hydrogel meshes. In contrast with non-degradable dex-MA hydrogels, degradable dex-lactate-HEMA gels with comparable network characteristics (degree of methacrylate substitution and initial water content) showed an almost zero-order, degradation controlled release of rhIL-2 in a time period of 5-15 days. This paper demonstrates that the release of rhIL-2 from non-degradable dex-MA and degradable dex-lactate-HEMA gels can be modulated by the crosslink density and/or the degradation characteristics of the hydrogel. Importantly, rhIL-2 was mainly released as monomer from the hydrogels and with good retention of its biological activity.

Dextrans↗
PubMed2001

A comparative biocompatibility study of microspheres based on crosslinked dextran or poly(lactic-co-glycolic)acid after subcutaneous injection in rats.

Microspheres based on methacrylated dextran (dex-MA), dextran derivatized with lactate-hydroxyethyl methacrylate (dex-lactate-HEMA) or derivatized with HEMA (dex-HEMA) were prepared. The microspheres were injected subcutaneously in rats and the effect of the particle size and network characteristics [initial water content and degree of methacrylate substitution (DS)] on the tissue reaction was investigated for 6 weeks. As a control, poly(lactic-co-glycolic)acid (PLGA) microspheres with varying sizes (unsized, smaller than 10 microm, smaller and larger than 20 microm) were injected as well. A mild tissue reaction to the PLGA microspheres was observed, characterized by infiltration of macrophages (MØs) and some granulocytes. Six weeks postinjection, the PLGA microspheres were still present. However, their size was decreased indicating degradation and many spheres had been phagocytosed. The tissue reaction was hardly affected by size differences, except for particles smaller than 10 microm, which induced an extensive tissue reaction. The initial tissue reaction to nondegradable dex-MA microspheres was stronger than towards the PLGA microspheres, but at day 10 the tissue reactions were comparable for both groups. Six weeks postinjection, the dex-MA microspheres were completely phagocytosed, and no signs of degradation were observed. The size and initial water content of dex-MA microspheres hardly affected the tissue response, although less granulocytes were observed for microspheres with higher DS. Slowly degrading dextran microspheres composed of dex-(lactate(1)-)HEMA induced a tissue reaction comparable to the PLGA microspheres. However, degradation of the dex-(lactate(1,3)-)HEMA microspheres was associated with an increased number of MØ's and giant cells, both phagocytosing the microspheres and their degradation products. Similar to PLGA, no adverse reactions were observed for the nondegradable dex-MA and degradable dextran microspheres. This study shows that both nondegradable and degradable dextran-based microspheres are well tolerated after subcutaneous injection in rats, which make them interesting candidates as controlled drug delivery systems.

Animals↗
PubMed2001

Oxidation of recombinant human interleukin-2 by potassium peroxodisulfate.

PURPOSE: The oxidation of recombinant human interleukin-2 (rhlL-2) by potassium peroxodisulfate (KPS) with or without N,N,N',N'-tetramethylethylenediamine (TEMED), which are used for the preparation of dextran-based hydrogels, was investigated. METHODS: The oxidation of (derivatives of) methionine. tryptophan, histidine and tyrosine, as well as rhlL-2 was investigated. Both the oxidation kinetics (RP-HPLC) and the nature of the oxidation products (mass spectrometry) were studied as a function of the KPS and TEMED concentration, and the presence of a competitive antioxidant, methionine. RESULTS: Under conditions relevant for the preparation of rhIL-2 loaded hydrogels, only methionine and tryptophan derivatives were susceptible to oxidation by KPS. The oxidation of these compounds was inhibited once TEMED was present, suggesting that the peroxodisulfate anion, rather than the radicals formed in the presence of TEMED, is the oxidative species. KPS only induced oxidation of the four methionines present in rhIL-2, whereas the tryptophan residue remained unaffected. The radicals, formed after KPS decomposition by TEMED, induced some dimerization of rhIL-2. The oxidation of rhIL-2 could be substantially reduced by the addition of methionine, or by pre-incubation of KPS with TEMED. CONCLUSIONS: Only the methionine residues in rhlL-2 are oxidized by KPS. The extent of oxidation can be minimized by a proper selection of the reaction conditions.

Electrophoresis, Polyacrylamide Gel↗
PubMed2000

In vivo biocompatibility of dextran-based hydrogels.

Dextran-based hydrogels were obtained by polymerization of aqueous solutions of methacrylated dextran (dex-MA) or lactate-hydroxyethyl methacrylate-derivatized dextran (dex-lactate-HEMA). Both nondegradable dex-MA and degradable dex-lactate-HEMA disk-shaped hydrogels, varying in initial water content and degree of substitution (DS, the number of methacrylate groups per 100 glucose units), were implanted subcutaneously in rats. The tissue reaction was evaluated over a period of 6 weeks. The initial foreign-body reaction to the dex-MA hydrogels was characterized by infiltration of granulocytes and macrophages and the formation of fibrin, and exudate, as well as new blood vessels. This reaction depended on the initial water content as well as on the DS of the hydrogel and decreased within 10 days. The mildest tissue response was observed for the gel with the highest water content and intermediate DS. At day 21 all dex-MA hydrogels were surrounded by a fibrous capsule and no toxic effects on the surrounding tissue were found. No signs of degradation were observed. The initial foreign-body reaction to the degradable dex-lactate-HEMA hydrogels was less severe compared with the dex-MA gels. In general, the size of the dex-lactate-HEMA hydrogels increased progressively with time and finally the gels completely dissolved. Degradation of the dex-lactate-HEMA hydrogels was associated with infiltration of macrophages and the formation of giant cells, both of which phagocytosed pieces of the hydrogel. A good correlation between the in vitro and the in vivo degradation time was found. This suggests that extra-cellular degradation is not caused by enzymes but depends only on hydrolysis of the ester and/or carbonate bonds present in the crosslinks of the hydrogels. After 21 days, the degradable hydrogels, as such, could not be retrieved, but accumulation of macrophages and giant cells was observed, both of which contained particles of the gels intracellularly. As for the dex-MA hydrogels, no toxic effects on the surrounding tissue were found. The results presented in this study demonstrate that dextran-based hydrogels can be considered as biocompatible materials, making these hydrogels attractive systems for drug delivery purposes.

Animals↗
PubMed1997

Analgesics in mice used in cancer research: reduction of discomfort?

During the last decades, an increase is apparent in the use of analgesics for laboratory animals in situations where this was previously considered unnecessary. Mice with advanced tumours often show clear signs of discomfort which may be a result of chronic pain or a result of general ill-being. The syngeneic murine tumour model most frequently used in our experiments was used to investigate whether this discomfort can be reduced with an analgesic. Twenty DBA/2 mice bearing SL2 lymphoma were given 0.5 mg/kg buprenorphine (Temgesic) in food gel twice daily, 20 tumour-bearing mice were given control food gel at the same times. Indicators of well-being were monitored daily. These included behavioural parameters such as exploration, grooming, and posture; food and water consumption and fur quality. All mice showed a clear increase of discomfort with time: explorative behaviours and grooming decreased, while sitting in hunched posture increased. Food and water consumption and fur quality also decreased. Major significant differences between the buprenorphine treated group and the control group were not apparent. In conclusion, we could not document a positive effect or buprenorphine on discomfort in mice as evaluated by our scoring system. It remains possible that pain itself was not the primary cause of the discomfort in mice bearing these tumours, or that the analgesic effect of buprenorphine was insufficient under these circumstances.

Analgesia↗
PubMed1994

Growth advantage of human leiomyoma cells compared to normal smooth-muscle cells due to enhanced sensitivity toward insulin-like growth factor I.

Human uterine leiomyomas exhibit increased IGF-I binding compared to myometrium, while both tissues show IGF-I gene expression. In this study we have examined the functional importance of these findings by testing the presence of IGF-I in 15 leiomyoma biopsies and in 18 myometrium biopsies and the capacity of smooth-muscle cells cultured from these tissues to react to IGF-I. The mean IGF-I peptide concentration in leiomyomas was 3 times higher than in myometrium. This resulted from increased IGF-I uptake in leiomyomas rather than from increased synthesis, as these tissues contain higher concentrations of type-I IGF receptors, as detected by immunohistochemistry, and equal levels of IGF-I mRNA. Blocking IGF-I transport with cytochalasin-B and with the type-I IGF receptor blocking antibody alpha IR3 in cultured cells induced decreased immunostaining intensity for IGF-I in most myometrium and leiomyoma cultures, indicating that the detected IGF-I is internalized. Depending on the culture conditions, IGF-I administration yielded increased survival or a higher proliferation rate in leiomyoma cultures than in myometrium cultures, indicating the increased importance of exogenous IGF-I for the growth of transformed smooth-muscle cells. We conclude that the increased concentrations of type-I IGF receptors in leiomyoma compared to myometrial smooth-muscle cells are functional with respect to the enhanced internalization of IGF-I and that they provide these tumor cells with a growth advantage compared to their normal counterparts.

Adult↗
PubMed1994

Effects of isoniazid (INH) on the BCG-induced local immune response after intravesical BCG therapy for superficial bladder cancer.

Because recent investigations showed that the use of isoniazid (INH) severely impaired the local immune reaction to intravesical bacillus Calmette-Guérin (BCG) in the bladder of guinea pigs, in this study the effect of INH in man has been investigated. Patients were treated with BCG with or without oral INH. The concentration of free INH in most urine samples of patients treated with BCG/INH was much higher (mean 38.0 +/- 60.9 micrograms INH/ml) than the minimal inhibitory concentration (MIC; 0.1 microgram INH/ml), suggesting at least a bacteriostatic potential of the INH present. However, in vitro studies showed that these urinary concentrations of INH did not kill BCG organisms effectively, even at a concentration of 150 micrograms/ml for 24 h. After the fifth and sixth BCG instillations a significant increase in the concentration of cytokines (IL2, IL6, IL8 and TNFa), IgG and IgA antibodies to BCG and the number of leukocytes in urine was observed. The leukocytes mainly consisted of granulocytes, besides monocytes/macrophages and, in lower amounts, T- and B-lymphocytes and natural killer (NK) cells. The absolute number of granulocytes and the concentration of IgG antibodies after BCG instillation were significantly suppressed by INH, whereas INH appeared to have no effect on the urinary cytokine and IgA antibody concentrations or the total number and phenotype of the leukocytes present. In conclusion, the results of this study indicate that INH does not impair the local immunological stimulation after BCG instillation in man as severely as was observed in the guinea pig and it may be expected that INH does not impair the antitumor efficacy of BCG.

Administration, Intravesical↗
PubMed1993

Suppression of human cartilage proteoglycan synthesis by rheumatoid synovial fluid mononuclear cells activated with mycobacterial 60-kd heat-shock protein.

OBJECTIVE: To examine whether T cell reactivity toward heat-shock proteins (HSP) contributes to cartilage destruction in rheumatoid arthritis (RA). METHODS: An in vitro system was used, in which human cartilage explants were cocultured with hsp60-activated synovial fluid mononuclear cells (SFMC) from patients with RA, and proteoglycan (PG) synthesis was measured. RESULTS: The hsp60-activated SFMC suppressed cartilage PG synthesis. This effect was dependent on the production of interleukin-1 (IL-1) and tumor necrosis factor alpha (TNF alpha). CONCLUSION: Mycobacterial 60-kd heat-shock protein can activate rheumatoid SFMC to suppress human cartilage PG synthesis. This suppression is mediated by IL-1 and TNF alpha.

Adult↗
PubMed1992

Expression of the first calcitonin/CGRP gene in spontaneous and transplanted rat medullary thyroid carcinoma: a comparison of dot-blot analysis, in situ hybridization, and immunohistochemistry.

Calcitonin (CT) and calcitonin gene-related peptide (CGRP) are encoded by a single gene, the CALC-I gene. They are expressed in the thyroid and in the nervous system by alternative splicing of the pre-messenger RNA derived from the CALC-I gene. In medullary thyroid carcinoma (MTC), a malignancy derived from the calcitonin-producing C-cells in the thyroid, production of calcitonin and CGRP is a common feature. We investigated the CT and CGRP production of four spontaneous MTCs transplanted three to four times and 14 MTC lines transplanted for several years in WAG/Rij rats, a strain with hereditary MTC. The expression of CT and CGRP in the spontaneous and in the transplanted tumors was studied by means of RNA in situ hybridization (RISH), dot-blot analysis, and immunohistochemistry. A down-regulation of CT production in transplanted compared with spontaneous tumors was observed, but an inverse relation between CT and CGRP mRNA content in both spontaneous and transplanted tumors was not observed. In this study, RISH proved to be as sensitive as dot-blot analysis to detect gene expression in tissue samples. The different approaches of analyzing the gene expression in tissue samples (the cellular localization of gene expression by ISH vs the analysis of an extract of a total tissue sample with dot-blot analysis) showed that each technique is equal in value and that they are complementary to each other.

Animals↗
PubMed1991

Role of TNF alpha, in relation to IL-1 and IL-6 in the proteoglycan turnover of human articular cartilage.

In both young and old human articular cartilage explants, TNF alpha induced a concentration-dependent, reversible suppression of the proteoglycan (PG) synthesis. Young cartilage was more sensitive to TNF alpha than old cartilage: 50% suppression of PG synthesis was reached at a TNF alpha concentration of 5 U/ml for young and 30 U/ml for old cartilage, whereas at 10(3) U/ml the PG synthesis of young cartilage was blocked and that of old cartilage suppressed by 80%. These inhibition levels of PG synthesis resulted in 25% PG depletion of the explants after 8 days of culture. The release of cartilage PG was not enhanced. TNF alpha induced no detectable amounts of IL-1 (less than 0.01 U) in young or old cartilage but did induce IL-6 production. The induced amounts of IL-6 were higher in young than in old cartilage but no dose-dependency was evident. Antibodies to neither IL-1 nor IL-6 had any influence on the TNF alpha-induced suppression of PG synthesis. The combination of TNF alpha and IL-1 led to an additive inhibition of PG synthesis which had no relationship to induced IL-6. TNF alpha was about 100-fold less active than IL-1.

Aging↗
PubMed1990

Interleukin-1-induced interleukin-6 is required for the inhibition of proteoglycan synthesis by interleukin-1 in human articular cartilage.

Cartilage from normal controls, patients with osteoarthritis, and patients with rheumatoid arthritis produced no interleukin-6 (IL-6) in culture. However, IL-1 induced massive production of IL-6 (up to 135 ng/ml) in cartilage from all 3 sources, in a dose-dependent manner (in some cases, a peak value was reached). The levels of induced IL-6 were similar to those found in rheumatoid arthritis synovial fluid. At IL-1 concentrations that induced almost complete inhibition of proteoglycan (PG) synthesis, IL-6 production could still be increased considerably. Exogenous IL-6 inhibited PG synthesis by up to 25%. IL-1-induced inhibition of PG synthesis was reversed by antibodies against recombinant human IL-6. These results suggest that IL-6 is required for the IL-1-induced inhibition of PG synthesis.

Adult↗
PubMed1988

Lack of correlation between in vivo peroxidatic activity patterns and tumoricidal activity in vitro of peritoneal macrophages after intraperitoneal immunization with tumor cells.

Immunization of C57BL mice with one inoculum of 10(7) DBA/2-derived SL2 lymphosarcoma cells resulted in a +/- 20-fold increase in the total number of peritoneal cells. The number of macrophages showed a 10-fold increase from 3 x 10(6) (control mice) to 3.4 x 10(7) cells at day 8 after immunization. Within this macrophage population, four different cell types, based on the ultrastructural peroxidatic activity patterns, could be distinguished: exudate macrophages, resident macrophages, resident-exudate macrophages and peroxidatic-activity-negative macrophages. The number of exudate macrophages significantly increased in the peritoneal cavity after immunization: at day 8 after immunization, a peak value of 10(7) cells was observed. At the same time, there were 2.2 x 10(7) peroxidase-activity-negative macrophages present (representing the control value x 50). Significant in vitro tumoricidal activity of the isolated macrophages could not be measured until 8 days after immunization. At that time, a cytotoxicity index of 68 was reached. After immunization of the C57BL mice with 3 injections with allogeneic SL2 cells, there were no dramatic changes in the number of peritoneal cells after the last immunization. Only immediately after the last immunization was a minor increase in peroxidatic-activity-negative macrophages seen. But already at 5 days after the last immunization, the composition of the peritoneal suspension was similar to that of non-immunized mice with predominantly resident macrophages. The cytotoxicity of the peritoneal macrophages from hyperimmunized mice was constantly high during 1-15 days after the last immunization (cytotoxicity index ranged from 66-72). In order to study which type(s) of macrophage(s) (resident, exudate, resident-exudate or peroxidatic-activity-negative) is/are responsible for the cytotoxicity measured in vitro, peritoneal cell suspensions (obtained after immunization) were fractionated according to their affinity to wheat germ agglutinin (WGA) coupled to Sepharose columns. Comparison of the values of cytotoxicity measured before and after separation into "subtypes" of the macrophages revealed that the expression of cytotoxicity is not correlated with any of the "sub-types", especially when the peroxidatic activity pattern is is taken as a criterion.

Animals↗
PubMed1987

Differential responses of old human cartilage explants to synovial- and mononuclear-cell factors.

To investigate mechanisms of cartilage destruction that may apply to rheumatoid arthritis, young and old human, and young porcine, articular cartilage was cultured for 8 days and the effects on proteoglycan (PG) metabolism of normal synovium supernatant (NSS), rheumatoid synovial fluid (RFL), and blood mononuclear cell supernatant (MCF) were studied. The effects were chondrocyte-mediated. An inverse correlation was found between baseline net PG synthesis and the effect of NSS on PG synthesis. Responses of young (porcine and human) cartilage were similar. In young cartilage the three agents induced PG depletion by suppression of net PG synthesis. In old cartilage NSS and RFL induced PG depletion, whereas MCF did not. In cartilage of low baseline net PG synthesis, NSS and MCF stimulated both PG release and PG synthesis; NSS stimulated predominantly PG release, and MCF predominantly PG synthesis. In conclusion, young and old human cartilage differ in the quality of their in vitro response to potentially catabolic factors. This may be due to the difference in baseline net PG synthesis. Synovial extracts differ from mononuclear-cell supernatants in their effects on old cartilage. It is suggested that this is caused by the presence, in different relative amounts, of factors that influence either PG synthesis or PG release.

Aging↗
PubMed1987

Carcinogenesis revisited.

The importance of mutations in carcinogenesis is still unclear. Assuming that mutations of the genetic material are central to this problem, the number needed to give rise to a cancer cell must be established. A one-mutation theory is unsatisfactory for a number of reasons. A four-mutation model fits better and can be calculated in humans assuming that the observed endogenous tumors in 2% of the population equals the frequency of spontaneous carcinogenesis, accepting a mean mutation rate of 2 X 10(-5) mutations per gene per generation, and a production of about 7 X 10(15) cells during our whole lifetime. This model is also consistent with the observed peak incidence of cancer in children, with the hereditary aspects of some pediatric tumors, and with the usually nonhereditary mechanisms of cancer in adults.

Humans↗
PubMed1986

Lymphocyte-induced macrophage cytotoxicity. III. Induction of specific macrophage cytotoxicity is independent of lipopolysaccharide.

The induction of specific macrophage cytotoxicity by allo-sensitized T cells in vitro is shown to be independent of the presence of lipopolysaccharide (LPS). This contrasts with the induction of macrophage cytotoxicity by Macrophage Activating Factor (MAF). The specific macrophage cytotoxicity could be induced in LPS-free medium (less than l ng/ml). Addition of LPS to the macrophages did not increase the cytotoxicity. Addition of LPS-binding polymyxin B to the macrophages before and during the induction of cytotoxicity did not reduce the specific macrophage cytotoxicity. Macrophages obtained from the LPS-unresponsive mouse strain C3H/HeJ were rendered cytotoxic by the allo-sensitized lymphocytes to the same extent as the macrophages from LPS-responsive C3HeB/Fe and C57BL mice. This indicates that the induction of macrophage cytotoxicity by MAF is different from the induction of specific macrophage cytotoxicity by Specific Macrophage Arming Factor (SMAF).

Animals↗